Researchers turn bacterial colonies into logic gates

Starting with a NOR gate created with DNA, researchers get colonies of …

For the past several years, researchers have been creating living logic gates, using the genetic on and off switches we've discovered in the biochemical world to create simple logical functions. But these systems typically run into a wall: we can only stuff so many functions into a single bacterium before the noisy processes within the cell start to interfere with their function, causing unpredictable results. Now, researchers have avoided this problem by getting small populations of bacteria to produce soluble factors that act as circuit wires. Arranged appropriately, these bacteria can perform any possible logical function.

The key to the new work is stretches of DNA that act as logical OR and NOR functions. Both of them rely on small stretches of DNA called promoters that control the activity of nearby genes. In this case, the authors used promoters that activate nearby genes in response to simple chemicals (arabinose and tetracycline for these two promoters). By putting both promoters next to a reporter gene, the system acted as an OR gate: when either of the chemicals was present, the reporter was on.

Promotors that are regulated by chemicals can be used to create an OR gate (left) or, with an extra step, a NOR gate.

To create a NOR gate, another layer of genetic circuitry was required. Instead of putting the two promoters next to the gene of interest, the authors linked them up to the gene for a protein that shuts down genes near any promoters it binds to (termed a repressor). Now, when either chemical was present, the repressor was expressed. They then linked their reporter gene to a promoter that was bound by the repressor. This inverted the final output: when either chemical was present, the reporter was shut down.

Because this is a logic gate, the same rules apply here as in electronic form, in that it's possible to build combinations of NOR gates that perform any logical function (AND, EQUAL, etc.). Researchers have typically done this by stuffing more combinations of promoters and regulators into a single bacterium, which, as we noted above, often runs into difficulties. So the team here did what we do with electronics: they wired bacteria together.

These aren't wires in the traditional sense. Instead, they relied on genes that bacteria use to signal each other. When produced by one bacterium, these quorum signals diffuse to its neighbors, where they regulate genes that change in response to the population density. In the recipient cell, they act very much like the chemicals used to control the logic gates, in that they link up with proteins that regulate promoters.

To turn these signals into wires, the authors simply replaced parts of their existing system. Instead of regulating a reporter gene, the NOR gate could be used to control the expression of one of these quorum signals. In the receiving cell, the promoters that respond to arabinose or tetracycline could be swapped out for a quorum-responsive promoter. Now, instead of responding to whether the authors put a chemical in the petri dish, some of the bacteria would respond to the output of their neighbors logical gates. As a result, different logic gates could be wired into a larger system.

Genetic logic gates can be used to regulate genes for quorum factors, which act as wires to transmit the logic results to other cells.

How does this possibly operate on a practical level? In this case, the authors set up small clusters of bacterial colonies (small lumps of genetically identical cells). Each colony had a single logic gate (the authors used NOR, OR, and NOT gates). Depending on the arrangement of the colonies, each one could signal to only one or two neighbors, and each could only take input from one or two. The authors demonstrated a functional XOR gate built from four colonies, showing that all logical functions can be built from similar combinations.

The nice thing about using populations of cells is that this averages out some of the chaotic behavior typical of systems based on single cells. At a minimum, the systems they tested showed a five-fold difference between their on and off states. The downside is that, relative to a single cell, these systems are huge. The authors suggest that it might be possible to adapt their system to single cells, but it's not clear that the same sort of performance could be maintained.

The other thing that would be nice to see is more complicated circuitry. It should be possible to link a few of these things up so that the final output is the product of several independent logic gates. Apparently, that will have to wait until the next paper.

Right now, these sorts of systems remain limited to labs; nobody has managed to find an application for a genetic circuit more complex than a simple on/off switch. But engineered bacteria are now being used for the production of everything from biofuels to pharmaceuticals, and there's a good chance that fine-grained control of cell behavior could be useful for some of these applications.